Photoactivatable Large Stokes Shift Fluorophores for Multicolor Nanoscopy

Area of Science:

• Chemical Biology
• Microscopy
• Organic Chemistry

Background:

• Photoactivatable probes are crucial for advanced cellular imaging.
• Existing probes often require caging groups or have limited spectral properties.

Purpose of the Study:

• To design and synthesize novel caging-group-free photoactivatable fluorescent dyes.
• To achieve red fluorescence emission and large Stokes shifts for improved imaging.
• To demonstrate compatibility with various microscopy techniques and labeling strategies.

Main Methods:

• Synthesis of novel 1-vinyl-10-silaxanthone imine derivatives.
• Characterization of photophysical properties, including fluorescence emission and Stokes shift.
• Application in multicolor fluorescence microscopy (fixed and live cells).
• Testing compatibility with super-resolution microscopy (STED, PALM).
• Demonstration of bioorthogonal labeling with tetrazine ligation and protein tags (HaloTag, SNAP-tag).

Main Results:

• Developed caging-group-free photoactivatable dyes with red fluorescence emission.
• Achieved a large Stokes shift of approximately 100 nm.
• Demonstrated byproduct-free one- and two-photon activation.
• Successfully applied the dyes in multicolor fluorescence microscopy of fixed and live cells.
• Validated compatibility with STED and PALM super-resolution techniques.
• Showcased utility in strain-promoted tetrazine ligation and self-labeling protein tag applications.

Conclusions:

• The novel silaxanthone imine-based dyes offer significant advantages for live-cell imaging.
• These probes facilitate advanced microscopy techniques, including super-resolution and multicolor imaging.
• The caging-group-free design and versatile labeling capabilities expand the toolkit for chemical biology research.